Pamela Tucker is trained in Infectious Diseases and has spent three years investigating the pathogenesis of acute alphavirus encephalitis. During this time she has acquired the foundation for a career in viral pathogenesis research. However, to ensure her success as an independent investigator, it is imperative that she secure protected time to continue developing her research potential. Alphaviruses are important causes of epidemic encephalitis in the Americas, yet the basic parameters of neurovirulence are unknown. Sindbis virus (SV)- the prototype alphavirus - causes age-dependent encephalitis in ice and provides an excellent animal model for studying the mechanisms of neurovirulence. SV has three structural proteins (C, E1, E@). Study of recombinant viruses has shown that a limited number of amino acid charges in the E1 and E2 surface glycoproteins are important determinants. Initial studies have shown that amino acid at E2-172 (Gly or Arg) affects replication in and binding to neuronal cells. The amino acid ar E2-55 (His to Gln) is important for viral replication in differentiated neurons. Viruses with His at E2-55 are fatal for 1, 7 and 14-day old mice, while viruses with Gln are virulent only for 1 day-old mice. Previous mortality studies suggest that a change from Thr to Ile at E2-3 is attenuating for newborn mice. To delineate the mechanisms by which amino acid changes in the E2 glycoprotein affect neurovirulence, studies will be performed using recombinant or mutagenized of SV differing at positions 172, 55 and 3. These strains will be characterized by determining mortality in mice, growth in, binding to and entry into neuronal cells, differentiated and undifferentiated neural cells and cultured dorsal root ganglia neurons. Using a Mab that recognizes a binding site on SV for neuronal cells, the influence that these amino acid changes have on neutralization, and cell binding will be determined. This work will be performed in the Neurovirology Laboratory under the sponsorship of Dr. Diane Griffin. Dr. Griffin's laboratory includes five postdoctoral fellows, a graduate student and a visiting scientists. The Neurovirology is a multidisciplinary training environment that includes 4 other independent investigators working on various aspects of the pathogenesis of infection with herpes simplex, Epstein Barr, measles, human and feline Immunodeficiency viruses.